PRISM hepatitis B surface antigen detection of hepatits B virus minipool nucleic acid testing yield samples

BACKGROUND: Hepatitis B virus (HBV) residual risk has been estimated at 1:63,000-1:205,000 and introduction of more sensitive serological tests and nucleic acid testing (NAT) would reduce that risk. Sensitivity of the recently licensed Abbott PRISM hepatitis B surface antigen (HBsAg) CLIA and minipool (MP) HBV NAT has been described as comparable and thus the need for HBV NAT has not been compelling. In this study, eight samples identified as yield samples with MP HBV NAT were tested using the PRISM test. STUDY DESIGN AND METHODS: Seven samples were identified using the Roche COBAS AmpliScreen HBV test and one additional sample was obtained from the clinical trial for the Roche cobas TaqScreen MPX test. Each of these samples was reactive by MP HBV NAT and nonreactive for HBsAg using one of three licensed enzyme immunoassay (EIA) tests. After licensure of the PRISM HBsAg, aliquots were tested with this assay, and DNA quantitation and genotyping were repeated where sample volume permitted. RESULTS: Three samples (2000, 2300, and 61,000 copies/mL) produced reactive results with PRISM. Four samples with viral loads less than 300 copies per mL produced nonreactive results. One sample, originally quantitated at 37,000 copies per mL (but 3850 copies/mL in repeat testing) was also nonreactive by PRISM. Genotyping of this sample indicated a type C genotype with no mutations. CONCLUSION: Adding serological sensitivity of PRISM CLIA reduced the NAT yield from the original 1: 385,555 to 1:610,488. However, MP HBV NAT still provides additional sensitivity over CLIA, even for a donation with a viral load of almost 4000 copies per mL.     

核酸扩增检测技术在血液筛查中的应用

目的探讨核酸扩增检测技术在血液筛查中的应用价值。方法用酶联免疫吸附试验(ELISA)常规筛查血液,对免疫指标合格的血液采用美国Roche Cobas Amplicor全自动PCR诊断系统检测HBV DNA、HCV RNA和HIV-1 RNA,样本混合采用48人份×50μL汇集,采用汇集池阳性的再分拆检测。结果 70 953份无偿献血标本中,HBV DNA阳性10份(1.4/10 000),未发现HCV RNA和HIV-1 RNA阳性样本。结论血站系统采用微量标本混合方式使用核酸扩增检测技术筛查血液是可行的,能有效提高临床用血的安全。     

A comparison of human immunodeficiency virus,hepatitis C virus,hepatitis B virus,and human T‐lymphotropic virus marker rates for directed versus volunteer blood donations to the American Red Cross during 2005 to 2010

BACKGROUND: At most US blood centers, patients may still opt to choose specific donors to give blood for their anticipated transfusion needs. However, there is little evidence of improved safety with directed donation when compared to volunteer donation. STUDY DESIGN AND METHODS: The percentage of directed donations made to the American Red Cross (ARC) from 1995 to 2010 was determined. Infectious disease marker rates for human immunodeficiency virus (HIV), hepatitis C virus (HCV), hepatitis B virus (HBV), and human T‐lymphotropic virus (HTLV) were calculated for volunteer and directed donations made from 2005 to 2010. Odds ratios (ORs) were calculated to compare marker‐positive rates of directed donations to volunteer donations. RESULTS: The percentage of donations from directed donors declined from 1.6% in 1995 to 0.12% in 2010. From 2005 to 2010, the ARC collected 38,894,782 volunteer and 69,869 directed donations. Rates of HIV, HCV, HBV, and HTLV for volunteer donations were 2.9, 32.2, 12.4, and 2.5 per 100,000 donations, respectively; for directed, the rates were 7.2, 93.0, 40.1, and 18.6 per 100,000. After demographics and first‐time or repeat status were adjusted for, corresponding ORs of viral marker positivity in directed versus volunteer donations were not significant for HIV, HBV, or HTLV and significant for HCV (OR, 0.7; 95% confidence interval, 0.50‐0.90). CONCLUSIONS: Directed donations have declined by 92% at the ARC since 1995, but have higher viral marker rates than volunteer donations. The difference can be explained in part by the effects of first‐time or repeat status of the donors. Patients considering directed donation should be appropriately counseled about the potential risks.     

献血者血液乙型肝炎病毒表面抗原确认试验

目的 对使用国产HBsAg酶联免疫吸附试验(ELISA)一步法试剂盒检测无偿献血者血液(包括机采血小板献血体检者)的可靠性进行评价.方法 对无偿献血者血液样品,使用国产的HBsAg酶联免疫吸附试验(ELISA)试剂盒进行测定.结果 呈阳性反应者、3种试剂检测结果不一致的部分可疑结果者和3种试剂检测为阴性者,使用珠海丽珠试剂公司研发的乙型肝炎病毒表面抗原(HBsAg)确认试剂盒(中和试验法) 初步进行确认试验.确认试验方法参照丽珠公司提供的说明书进行.结果 132例样品中阳性反应或可疑阳性反应者123例被确认为阳性者106例,其中有17例可疑结果者常规确认试验结果为"阴性".经3种试剂再测,结果仍为可疑.9例0.074≤样品A值＜Cut off的样品和随机抽取的9例阴性样品常规确认试验结果仍为阴性.8例单一试剂呈阳性反应者常规确认为假阳性.结论 国产HBsAg ELISA试剂测定结果为阳性、弱阳性和可疑的样品应使用适当方法进行确认,排除假阳性,以保证结果准确.目前国产HBsAg酶联免疫吸附试验(ELISA)一步法试剂盒有较高的灵敏度,特异性尚待提高.     

Sensitivity of hepatitis B virus DNA transcription-mediated amplification testing in hepatitis B surface antigen-positive blood donations

BACKGROUND: The objective was to evaluate the performance of nucleic acid testing (NAT) in the detection of hepatitis B virus (HBV) infection in hepatitis B surface antigen (HBsAg)-positive blood donations. STUDY DESIGN AND METHODS: A total of 253 HBsAg- and anti-hepatitis B core antigen (HBc)-positive samples (50 hepatitis B e antigen [HBeAg]-positive and 203 anti-HBe-positive) from blood donations collected in France were studied. The samples were investigated with a blood screening assay (Procleix Ultrio, Chiron/Gen-Probe) in minipool (MP; x8) and in individual-donation (ID) testing. All nonreactive samples were retested once, and nonreactive MP samples were assayed for viral load (VL). RESULTS: All 50 HBeAg-positive samples were reactive in MP-NAT and ID-NAT. Of the 203 anti-HBe-positive donations, 80.3 percent were MP- and ID-reactive, 17.2 percent were MP-nonreactive and ID-reactive, and 2.5 percent were nonreactive in ID-NAT. Overall the sensitivity of ID-NAT was 98 percent versus 84 percent for MP-NAT. After retesting, 16 of the 35 MP-nonreactive and/or ID-reactive donations became MP-reactive and 2 of the ID-nonreactive donations became NAT-reactive. The capacity of Procleix Ultrio to detect HBV DNA was not related to HBsAg subtype, but correlated with the VL: the mean VL in the group of MP-nonreactive samples was 1,420 copies per mL vs. 17,000 copies per mL in the group of 40 MP-reactive samples. CONCLUSION: These results demonstrate that HBV-NAT in ID format is far more effective in detecting viremia in chronic HBsAg carriers than in MP-NAT. The sensitivity of the NAT assay needs to be improved to be considered for replacing the current HBsAg assays, especially when anti-HBc testing is not performed.     

Hepatitis B virus (HBV) DNA screening of blood donations in minipools with the COBAS AmpliScreen HBV test

BACKGROUND: The risk of hepatitis B virus (HBV) transmission by blood transfusion (estimated at 1 in 63,000-1 in 205,000 units in the United States) exceeds that of hepatitis C virus (HCV) or human immunodeficiency virus (HIV). Reduction of window-period HBV transmissions through detection of HBV DNA-positive units by minipool nucleic acid testing (MP NAT) would be expected to decrease this risk. STUDY DESIGN AND METHODS: A large multicenter study of the COBAS AmpliScreen HBV test (Roche Molecular Systems) was conducted on minipools of 24 blood donation specimens. The yield of HBV DNA-positive, hepatitis B surface antigen (HBsAg)-negative window-period donations was determined relative to current and newly licensed HBsAg assays. Donors with selected HBV DNA, HBsAg, and anti-hepatitis B core antigen (HBc) results were further evaluated. RESULTS: The detection rate of window-period units was 1 in 352,451 (95% confidence interval, 1 in 2,941,176-1 in 97,561). Assay specificity was high (99.9964%). HBV DNA was detected in 84 percent of HBsAg-positive, anti-HBc-positive donations by MP NAT and in 94 percent when individual-donation (ID) NAT was added. HBV DNA was detected in 0.03 percent of HBsAg-negative, anti-HBc-positive donations by MP NAT and in 0.41 percent when ID NAT was added. CONCLUSIONS: Implementation of HBV MP NAT will provide an increment in safety relative to HBV serologic screening, similar to that for HCV and in excess of that for HIV. Our data indicate that the implementation of HBV MP NAT would likely interdict 39 HBV window-period units and prevent 56 cases of transfusion-transmitted HBV infection annually. The current data indicate that HBV MP NAT should not lead to discontinuation of anti-HBc testing but that discontinuation of HBsAg testing with retention of anti-HBc testing may be possible.     

Two cases of transfusion‐transmitted hepatitis B virus (HBV) infection in a low‐endemic country before implementation of HBV nucleic acid testing

BACKGROUND: The risk of hepatitis B virus (HBV) transmission by transfusion is higher than that of other blood‐borne viruses. In France, before the introduction of HBV nucleic acid testing (NAT) in 2010, blood donations were tested for hepatitis B surface antigen (HBsAg) and antibodies against hepatitis B core antigen, and the residual risk of HBV transfusion related to preseroconversion acute phase was estimated at 0.54 per million donations. The additional value of the implementation of a highly sensitive HBV NAT to prevent such transmissions is discussed. STUDY DESIGN AND METHODS: Two lookback investigations based on HBV seroconversion of repeat donors were performed. Donors and recipients were followed up in multiple samples that were tested for HBV serologic and molecular markers. RESULTS: The recipients have shown posttransfusion HBsAg seroconversion. The archived samples from the implicated donations were positive for HBV DNA at extremely low viral load in both cases. HBV isolates from donors and recipients of each case were organized in the same cluster with 100% identities into Genotypes A2 and B4, respectively. One recipient spontaneously recovered from infection while the second was successfully treated. CONCLUSION: The present cases highlight the importance of introducing highly sensitive HBV NAT to prevent transmission. Moreover, the lookback studies based on appropriate molecular and serologic investigations of patients transfused with previous donations from newly identified HBV‐infected repeat donors offer the opportunity to treat a recently infected recipient.     

Cost-effectiveness of additional hepatitis B virus nucleic acid testing of individual donations or minipools of six donations in the Netherlands

BACKGROUND: To further reduce the risk of hepatitis B virus (HBV) transmission by blood transfusion, nucleic acid testing (NAT) can be employed. The aim of this study is to estimate the incremental cost-effectiveness ratio (ICER) in the Netherlands of employing a triplex NAT assay aimed at HBV nucleic acid detection in individual donations (ID-NAT) or in minipools of 6 donations (MP-6-NAT), compared to a triplex NAT assay in minipools of 24 donations (MP-24-NAT).
STUDY DESIGN AND METHODS: A mathematical model was made of the whole transfusion chain from donors to recipients of blood in the Netherlands. The annual number of avoided HBV transmissions was estimated with the window-period incidence model. The natural history of a HBV infection in recipients is described by a Markov model.
RESULTS: The ICER of adding HBV MP-6-NAT or HBV ID-NAT in the Netherlands is €303,218 (95% confidence interval [CI], €233,001-€408,388) and €518,995 (95% CI, €399,359-€699,120) per quality-adjusted life-year, respectively. The ICER strongly correlates with the age of transfusion recipients.
CONCLUSION: The cost-effectiveness of additional HBV NAT is limited by the limited loss of life caused by HBV transmission. Despite a higher effectiveness, HBV ID-NAT is less cost-effective than MP-6-NAT due to higher costs. A future equivalent participation of immigrants from HBV-endemic countries in the donor base renders HBV NAT only slightly more cost-effective.     

核酸扩增检测在血液筛查的初步应用

目的为提高血站供血安全性,探讨核酸扩增检测在血站血液筛查中的可行性。方法在酶联免疫吸附试验(ELISA)常规筛查血液的基础上,采用荧光定量聚合酶链反应（PCR）方法,检测乙型肝炎表面抗原(HBsAg)、丙型肝炎抗体(抗-HCV)和人免疫缺陷病毒抗体(抗-HIV)1/2呈阴性的献血者微量血浆汇集池标本（20人份×50μl汇集）中的丙肝病毒（HCV）和乙肝病毒（HBV）核酸,再对阳性汇集池中的标本进行单份检测。结果8805份（分442个汇集池）血液被检测HCVRNA,结果有1例（0.01%）为阳性;1441份血液被检测HBVDNA,结果6例（0.4%）为阳性。从标本汇集到筛查出单份阳性献血者大约需要3d。结论ELISA结合荧光定量PCR检测微量血浆汇集池标本的方法筛查血液HBV和HCV感染是可行的,能够进一步提高输血的安全性。     

A novel hepatitis B virus surface antigen immunoassay as sensitive as hepatitis B virus nucleic acid testing in detecting early infection

BACKGROUND: The aim was to considerably enhance the sensitivity of hepatitis B virus (HBV) surface antigen (HBsAg) detection and investigate whether the window period for HBV detection could be reduced.
STUDY DESIGN AND METHODS: A high-sensitivity chemiluminescent enzyme immunoassay (CLEIA) was developed for quantitative HBsAg detection by a combination of monoclonal antibodies, each one for a specific epitope of HBsAg, and by improving the conjugation technique. The sensitivity of the assay was compared with that of the existing chemiluminescent immunoassay (CLIA). Commercially available seroconversion panels and samples of HBV-infected chimpanzees were tested with the developed prototype to assess whether the window period for HBsAg detection could be reduced to that for DNA detection.
RESULTS: Compared to the existing CLIA, the CLEIA prototype detected HBsAg with approximately 230-fold higher sensitivity and showed a reduced window period. HBsAg detection by the CLEIA prototype and HBV DNA detection by polymerase chain reaction (PCR) occurred simultaneously. The mean time for the CLEIA prototype to first detect HBsAg was approximately 17.4 days less than that for the existing systems. Further, CLEIA prototype enabled HBsAg detection even in anti-HBs–positive seroconversion samples. In the inoculated chimpanzees the HBsAg and HBV DNA became detectable simultaneously and concentrations increased in parallel, whereas HBsAg remained detectable longer than HBV DNA in the declining phase of viremia.
CONCLUSION: The CLEIA prototype yielded results comparable with those of HBV DNA PCR. This novel high-sensitivity assay may be useful for early detection of HBV infection and monitoring patients with a history of infection.     

甲型流感病毒抗原及核酸检测联合应用研究

目的:通过大样本流感样病例检测,明确抗原检测与核酸检测两种方法联合应用原则,从而快速明确流感诊断,控制流感暴发,减少资源浪费。方法:采集2019年1月至2020年1月北京同仁医院发热门诊的4 622份流感样病例咽拭子标本,其中流感季3 230份,非流感季1 392份,采用快速抗原法和实时荧光逆转录聚合酶链反应（qPCR...     

One-year experience of nucleic acid technology testing for human immunodeficiency virus Type 1, hepatitis C virus, and hepatitis B virus in Thai blood donations

BACKGROUND: Blood donations collected at the National Blood Center, the Thai Red Cross Society, Bangkok, in 2007 were tested by nucleic acid amplification technology (NAT) using the Chiron TIGRIS/Procleix Ultrio test and the Roche cobas s 201/cobas TaqScreen multiplex (MPX) test.
STUDY DESIGN AND METHODS: The sensitivity, specificity, and robustness were determined by testing 486,676 seronegative blood donations. Samples from each day of collection were divided into two sets; the odd-numbered samples were tested individually on the TIGRIS and the even-numbered samples were tested in pools of 6 on the cobas s 201. The status of reactive samples was confirmed by duplicate testing of samples from the plasma bag to calculate the test specificity. Reactive samples were tested on the alternate system and followed up.
RESULTS: The analytical sensitivity of both systems met the 95% limits of detection claimed by the respective package inserts. No cross contamination was seen with either system. Test specificity was 99.93 and 99.90% for the Procleix Ultrio and cobas TaqScreen tests, respectively. The NAT yield rates for human immunodeficiency virus Type 1 (HIV-1), hepatitis C virus (HCV), and hepatitis B virus (HBV) were 1:97,000, 1:490,000, and 1:2800, respectively. Several occult HBV donors, the majority of whom were detected by both tests, were also identified. The HIV-1 and HCV window cases were detected with both tests.
CONCLUSION: The performances of the systems and tests indicated that both were acceptable for routine NAT by the National Blood Center, the Thai Red Cross Society. However, the Procleix Ultrio test appeared to be less sensitive than the cobas TaqScreen test for HBV.